Our research plan is focused on generating a comprehensive understanding of G-protein-coupled receptors (GPCRs) and their signaling partners on epithelial stem cell proliferation and differentiation. Our previous findings indicate that GPCR regulation of cAMP signaling may be involved in differentiation and tumorigenicity within the epidermal stem cell compartment. By using the skin as a somatic stem cell model system, we have characterized the GPCRs expressed in human keratinocytes under basal and differentiated conditions and find that G alpha i-coupled GPCRs can affect keratinocyte proliferation. We then used a chemogenetic approach to demonstrate that GPCR-G alpha i activation can regulate keratinocyte proliferation and differentiation and that overactivation of G alpha i-signaling in the basal and stem cell compartment of the skin can lead to epidermal hyperplasia. Our results expand our understanding of the role of GPCR-cAMP signaling in skin homeostasis and reveal overlapping and divergent roles of the cAMP-regulating heterotrimeric G alpha s and G alpha i proteins in stem cells. We are currently characterizing further the function of different GPCRs in epithelial cells. Among the downstream signaling pathways regulated by GPCRs, we are investigating the role of YAP1/TAZ signaling in regulating epithelial stem cell fate. By using a newly developed genetically-encoded inhibitor of YAP1/TAZ-mediated transcription, we have identified the gene networks regulated by this signaling pathway in human skin by utilizing normal and cancer cell lines as well as a newly established animal model. The YAP1/TAZ inhibitor is allowing us to determine the precise role of YAP1/TAZ in normal cell homeostasis and in cancer initiation and progression. In addition, we will be able to test the role of YAP1/TAZ/TEAD in transducing mitogenic signals and other transcriptional events initiated by GPCR signaling.